Plants orient the growth of their primary organs according to the gravity vector. In roots, a gravity stimulus induces a reorientation of growth in the elongation zone, provided the root cap is present. Various physiological events are triggered by a gravity stimulus in the root cap, including amyloplast sedimentation, changes in the patterns of electrical field surrounding the root tip and a lateral redistribution of Ca2+ and IAA. The long term objective of our research is to define the succession of molecular events involved in gravity sensing and signal transduction by the root cap of Arabidopsis thaliana. As a first step towards that objective, two insertional mutagenesis strategies will be used to identify genes expressed specifically in the root cap which are essential for root gravitropism. The Dupont de Nemours collection of T-DNA-insertional mutants (8,000 independent lines) will be screened for root agravitropic mutants. Lines showing an abnormal root gravitropism will be recovered, propagated by self- pollination, and characterized genetically. The tagged genes will be cloned and characterized, and their patterns of expression will be determined. Mutants affected in genes expressed in the root cap will be retained for further analysis. The amino acid sequence of the corresponding proteins will be deduced from the nucleic acid sequences and analyzed. The histological structure of the root cap in mutant plants will be compared to similar structures in wild-type plants. Similarly, auxin synthesis and transport, as well as the rate of auxin and Ca2+ redistribution in the root tip upon gravistimulation will be determined in mutant plants and compared to similar characteristics in wild-type plants. These genetic, cytological and physiological analyses of the mutants will allow the characterization of the functions of the tagged genes and, in turn, the cells expressing them. Similarly, "enhancer/promoter trap" systems using T-DNA sequences modified to contain a reporter gene whose expression is dependent on the action of the control sequences of the plant gene in which they are inserted will be developed and used to tag genes expressed specifically in the root cap of Arabidopsis thaliana. Transgenic plants containing insertions in such genes will be self-pollinated and the corresponding progeny will be tested for root gravitropism. Lines showing an abnormal root gravitropic response will be recovered and analyzed as described above. In the long term, the functional characterization of several root agravitropic mutants will provide the basis necessary to decipher the succession of events occurring in the root cap during gravistimulation. This project is also susceptible to provide a better understanding of several fundamental physiological and molecular processes involved in plant growth, development and response to environmental stimuli, including ion transport, changes in the patterns of electric currents, IAA transport and cell elongation.